My research objective is to elucidate the mechanism by which vitamin Bl2 transport proteins facilitate the cellular uptake of vitamin Bl2. Plasma transcobalamin II facilitates the uptake of vitamin Bl2 by a number of different cells. We have isolated human and rabbit transcobalmin II in homogeneous form and have developed methods for labeling these proteins with radioactive iodine. These labeled proteins will be used to study the role and fate of transcobalamin II during the complex process in which vitamin Bl2 binds to cell surfaces, enters cells, migrates to various sub-cellular fractions, and exits from cells. Co58-vitamin B12 and a variety of Co-57-viamin B12 analogues will be employed to study the importance of, and alterations which occur in, the structure of vitamin B12 during these processes. Human R-type vitamin B12-binding proteins with low sialic acid contents transport vitamin B12 exclusively to the liver. I-125-labeled R-type proteins, Co58-vitamin Bl2, and Co57-vitamin Bl2 analogues will be employed to elucidate this process and to determine its biological significance. We also plan to characterize the process by which structurally abnormal R-type proteins are synthesized in chronic myelogenous leukemia and other myeloproliferative diseases. Intrinsic factor facilitates the uptake of vitamin B12 by ileal mucosal cells. We plan to isolate and characterize the ileal cell surface receptor for the intrinsic factor-vitamin Bl2 complex. We plan to label intrinsic factor in vivo with H3-amino acids and utilize this radioactive protein to elucidate the mechanism by which intrinsic factor facilitates the ileal absorption of vitamin Bl2.